Identification of β-catenin-interacting proteins in nuclear fractions of native rat collecting duct cells

다국어 초록 (Multilingual Abstract)

<P>The gene encoding the aquaporin-2 water channel is regulated transcriptionally in response to vasopressin. In the renal collecting duct, vasopressin stimulates the nuclear translocation and phosphorylation (at Ser<SUP>552</SUP>) of β-catenin, a multifunctional protein that acts as a transcriptional coregulator in the nucleus. The purpose of this study was to identify β-catenin-interacting proteins that might be involved in transcriptional regulation in rat inner medullary collecting duct (IMCD) cells, using experimental and computational approaches. We used a standard chromatin immunoprecipitation procedure coupled to mass spectrometry (ChIP-MS) in a nuclear fraction isolated from rat IMCD suspensions. Over four biological replicates, we reproducibly identified 43 β-catenin-binding proteins, including several known β-catenin-binding partners as well as novel interacting proteins. Multiple proteins involved in transcriptional regulation were identified (Taf1, Jup, Tdrd3, Cdh1, Cenpj, and several histones). Many of the identified β-catenin-binding partners were found in prior studies to translocate to the nucleus in response to vasopressin. There was only one DNA-binding transcription factor (TF), specifically Taf1, part of the RNA-polymerase II preinitiation complex. To identify sequence-specific TFs that might interact with β-catenin, Bayes’ theorem was used to integrate data from several information sources. The analysis identified several TFs with potential binding sites in the <I>Aqp2</I> gene promoter that could interact with β-catenin in the regulation of <I>Aqp2</I> gene transcription, specifically Jun, Junb, Jund, Atf1, Atf2, Mef2d, Usf1, Max, Pou2f1, and Rxra. The findings provide information necessary for modeling the transcriptional response to vasopressin.</P>